Intestinal epithelial cells must detect and respond to microbial pathogens that cause foodborne and waterborne diseases, yet also discriminate them from the trillions of other microbes in the intestine that are innocuous or even beneficial. The mechanisms by which host cells make this distinction are poorly understood. Our long-term goal is to better understand how pathogens are discriminated from other microbes by intestinal cells, as well as by other cell types. Closing this gap in our understanding may allow for improved treatment of infectious diseases, as well as better control of inflammatory disorders. Our central hypothesis is that a major strategy for discriminating pathogens from other microbes is through sensing perturbations of core host processes that are commonly targeted by pathogen-derived toxins. The objective here is to identify the mechanism by which the nematode C. elegans detects such pathogen-induced perturbations. C. elegans provides an excellent system to address this question because it relies on epithelial defense and is extremely tractable. We will also extend our findings to mammals, to investigate a similar protective host response. Our recent findings have shown that perturbation of core processes like mRNA translation triggers activation of the C. elegans bZIP transcription factor ZIP-2 to provide host defense (Dunbar et al, 2012). ZIP-2 upregulates a suite of genes involved in intracellular defense, and promotes resistance to infection (Estes et al, 2010). Our unpublished results suggest that ZIP-2 works together with CEBP-2, which is the C. elegans ortholog of mammalian C/EBP-?, a transcription factor that mediates acute response to infection in mammals. We hypothesize that 1) ZIP-2 and CEBP-2 form a heterodimeric transcription factor that promotes host defense, 2) ZIP-2 translation is upregulated upon infection by an upstream open reading frame (uORF) that acts as a sensor for translational attenuation, and 3) mammals deploy a similar host defense system. We will test our first hypothesis in Specific Aim 1, where we will examine CEBP-2 and ZIP-2 phenotypes, expression and interaction. We will test our second hypothesis in Specific Aim 2, where we investigate the underlying mechanisms of the surprising result that translational attenuation increases ZIP-2 protein levels. We will analyze the zip-2 uORF (Dunbar et al, 2012), and characterize hits from a genetic screen for regulators of ZIP- 2. In Aim 3, we will build off our preliminary findings into mammals, testing a role for C/EBP-? in the protective response to toxins, and investigating the functional significance of this response in vivo. This approach is innovative, because it analyzes how cells detect perturbations in core processes targeted by pathogens, which is a newly appreciated mode of animal host defense. It also investigates the role and function of uORFs, which are found in 30-50% of all mouse and human genes, but poorly understood. The proposed research is significant because it will provide insight into how cells respond specifically to pathogenic attack, and may lead to new treatments for infections in the intestine as well as other tissues, to better combat infectious disease.